Calcium-dependent secretion constitutes the primary mode of communication in the nervous system. The long-term goal of this research is an understanding of the molecular events underlying this process. Modifications in neurotransmission affect many critical processes such as learning and memory, and perturbations in this system are thought to be involved in the etiology of several affective disorders. While significant progress has been made in this area over the last few years, many critical issues remain regarding the mechanisms that underlie and modulate secretion. The intracellular trafficking of neurotransmitter-containing vesicles is proposed to be mediated by the formation and dissolution of specific protein complexes composed of elements residing on vesicular and plasma membranes as well as cytoplasmic components. Hrs-2 is an ATPase that interacts with SNAP-25, plasma membrane-associated protein known to be critical for secretion. Hrs-2 is regulated by divalent cations known to influence secretion and inhibits secretion from permeabilized cells. We propose to further characterize the mechanism by which hrs-2 acts in the secretory process. A complementary, multi-faceted approach is being taken which includes: (1) analyzing hrs-2 protein interactions and their regulation, (2) defining the distribution and regulation of the membrane association of hrs-2, (3) examining the regulation of hrs-2 ATPase activity, and (4) characterizing the molecular mechanisms by which hrs-2 inhibits secretion.